Nakamura Jun, Tajima Genichi, Sato Chikara, Furukohri Takahiro, Konishi Kazuhiko
Department of Developmental Biology and Neurosciences, Graduate School of Life Sciences, Tohoku University, Aoba-ku, Sendai, Miyagi 980-8578, Japan.
J Biol Chem. 2002 Jul 5;277(27):24180-90. doi: 10.1074/jbc.M111834200. Epub 2002 Apr 25.
The effect of ATP on calcium binding of the Ca2+-ATPase of the sarcoplasmic reticulum has not been clarified. By comparing the calcium dependence of the ATPase activity and of phosphorylation of the ATPase molecules with that of calcium binding in the absence of ATP, we show the existence of two types of regulatory site of the enzyme molecules at which ATP binding variously improves the calcium binding performance of the molecules depending on the aggregation state of the molecules and pH; the two regulatory sites bind ATP at submillimolar (0.25 mm) and millimolar (5 mm) ATP, respectively. The results are discussed based on a model of two conformational variants (A and B forms) of the chemically equivalent ATPase molecules (Nakamura, J., and Furukohri, T. (1994) J. Biol. Chem. 269, 30818-30821). For example, in the sarcoplasmic reticulum membrane at pH 7.40, submillimolar ATP converted the calcium binding manner of the A form from noncooperative (Hill number (n(H)) of approximately 1) to cooperative (n(H) approximately 2), concurrent with a decrease in the apparent calcium affinity (K(0.5)) from 2-6 to 0.1-0.3 microm. The binding of the A form became almost the same as that of the B form (n(H) approximately 2, K(0.5) approximately 0.2 microm), which was not affected by ATP. Millimolar ATP further decreased the K(0.5) of the cooperative binding of the two forms to approximately 0.05 microm. Regulation of the calcium binding performance by ATP is discussed in terms of monomeric and oligomeric pathway models.
三磷酸腺苷（ATP）对肌浆网Ca2 + -ATP酶钙结合的影响尚未明确。通过比较在无ATP情况下ATP酶活性和ATP酶分子磷酸化的钙依赖性与钙结合的钙依赖性，我们发现该酶分子存在两种调节位点，ATP结合在这两个位点上会根据分子的聚集状态和pH值不同程度地改善分子的钙结合性能；这两个调节位点分别在亚毫摩尔（0.25 mM）和毫摩尔（5 mM）ATP浓度下结合ATP。基于化学等价的ATP酶分子的两种构象变体（A和B形式）模型（Nakamura, J., and Furukohri, T. (1994) J. Biol. Chem. 269, 30818 - 30821）对结果进行了讨论。例如，在pH 7.40的肌浆网膜中，亚毫摩尔ATP使A形式的钙结合方式从非协同性（希尔系数（n(H)）约为1）转变为协同性（n(H)约为2），同时表观钙亲和力（K(0.5)）从2 - 6 μM降至0.1 - 0.3 μM。A形式的结合变得与B形式几乎相同（n(H)约为2，K(0.5)约为0.2 μM），而B形式不受ATP影响。毫摩尔ATP进一步将两种形式协同结合的K(0.5)降低至约0.05 μM。从单体和寡聚体途径模型的角度讨论了ATP对钙结合性能的调节。
